Identification and role of the homodimerization interface of the glycosylphosphatidylinositol-anchored membrane type 6 matrix metalloproteinase (MMP25).
The membrane type (MT) 6 matrix metalloproteinase (MMP) (MMP25) is a glycosylphosphatidylinositol-anchored matrix metalloproteinase (MMP) that is highly expressed in leukocytes and in some cancer tissues. We previously showed that natural MT6-MMP is expressed on the cell surface as a major reduction-sensitive form of M(r) 120, likely representing enzyme homodimers held by disulfide bridges. Among the membrane type-MMPs, the stem region of MT6-MMP contains three cysteine residues at positions 530, 532, and 534 which may contribute to dimerization. A systematic site-directed mutagenesis study of the Cys residues in the stem region shows that Cys(532) is involved in MT6-MMP dimerization by forming an intermolecular disulfide bond. The mutagenesis data also suggest that Cys(530) and Cys(534) form an intramolecular disulfide bond. The experimental observations on cysteines were also investigated by computational studies of the stem peptide, which validate these proposals. Dimerization is not essential for transport of MT6-MMP to the cell surface, partitioning into lipid rafts or cleavage of alpha-1-proteinase inhibitor. However, monomeric forms of MT6-MMP exhibited enhanced autolysis and metalloprotease-dependent degradation. Collectively, these studies establish the stem region of MT6-MMP as the dimerization interface, an event whose outcome imparts protease stability to the protein.
Zhao
Huiren
H
Department of Pathology, Wayne State University and Proteases and Cancer Program, Barbara Ann Karmanos Cancer Institute, Detroit, Michigan 48201, USA.
Sohail
Anjum
A
Sun
Qing
Q
Shi
Qicun
Q
Kim
Seaho
S
Mobashery
Shahriar
S
Fridman
Rafael
R
D002454:Cell Differentiation; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D019295:Computational Biology / Q000379:methods; D003545:Cysteine / Q000737:chemistry; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D058851:GPI-Linked Proteins; D017261:Glycosylphosphatidylinositols / Q000737:chemistry*; D018922:HL-60 Cells; D006801:Humans; D008055:Lipids / Q000737:chemistry; D053504:Matrix Metalloproteinases, Membrane-Associated / Q000737:chemistry* / Q000378:metabolism; D021962:Membrane Microdomains; D016297:Mutagenesis, Site-Directed; D009504:Neutrophils / Q000378:metabolism
D004220:Disulfides; D058851:GPI-Linked Proteins; D017261:Glycosylphosphatidylinositols; D008055:Lipids; D053504:Matrix Metalloproteinases, Membrane-Associated; C403928:matrix metalloproteinase 25; D003545:Cysteine